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Title
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Studies of the domain structure of mammalian DNA polymerase beta. Identification of a discrete template binding domain.
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Authors
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A.Kumar,
S.G.Widen,
K.R.Williams,
P.Kedar,
R.L.Karpel,
S.H.Wilson.
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Ref.
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J Biol Chem, 1990,
265,
2124-2131.
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PubMed id
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Abstract
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Characterization of the domain structure of DNA polymerase beta is reported.
Large scale overproduction of the rat protein in Escherichia coli was achieved,
and the purified recombinant protein was verified by sequencing tryptic
peptides. This protein is both a single-stranded DNA binding protein and a DNA
polymerase consisting of one polypeptide chain of 334 amino acids. As revealed
by controlled proteolysis experiments, the protein is organized in two
relatively protease-resistant segments linked by a short protease-sensitive
region. One of these protease-resistant segments represents the NH2-terminal 20%
of the protein. This NH2-terminal domain (of about 75 residues) has strong
affinity for single-stranded nucleic acids. The other protease-resistant
segment, representing the COOH-terminal domain of approximately 250 residues,
does not bind to nucleic acids. Neither domain, tested as purified proteins, has
substantial DNA polymerase activity. The results suggest that the NH2-terminal
domain is principally responsible for the template binding activity of the
intact protein.
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